Use of bioreactors for large-scale multiplication of sugarcane (Saccharum spp.), energy cane (Saccharum spp.), and related species

The objective of this study was to set up a plant micropropagation facility to mass propagate sugarcane, energy cane, and related clonally propagated species. An efficient methodology for micropropagation of energy cane and perennial grasses using temporary immersion bioreactors was developed. Several different methods of tissue culture initiation, multiplication, and rooting were evaluated for several varieties of sugarcane (Saccharum officinarum L.) and sugarcane-related species such as Erianthus spp., Miscanthus spp., and Sorghum spp. × sugarcane hybrids, all from a germplasm collection. Apical meristem cultures were initiated for all genotypes that were micropropagated, when liquid or semisolid Murashige and Skoog (MS) medium was used, which was supplemented with 0.1–0.2 mg L−1 BAP, 0.1 mg L−1 kinetin, 0–0.1 mg L−1 NAA, and 0–0.2 μg L−1 giberellic acid. These cultures produced shoots between 4 and 8 wk after initiation. Shoot regeneration from leaf rolls or immature inflorescences was observed as early as 4 wk after initiation. Shoot multiplication was successful for all genotypes cultured in MS medium with 0.2 mg L−1 BAP and 0.1 mg L−1 kinetin. Energy cane had a significantly higher combined multiplication rate when grown under four or five LED lamps than when grown under three LED lamps, or under fluorescent lights in a growth chamber. The addition of 2 mg L−1 NAA produced faster and better rooting in all of the genotypes tested. Shoots produced well-developed roots after one cycle of 15–21 d in the bioreactors. The maximum number of plantlets produced per bioreactor was 1080. Plantlets developed a vigorous root system and were ready to be transplanted into the field after 2 mo. A protocol was standardized for different energy cane clones that were recommended for their biomass production and cell wall composition. Different tissues were used to speed up or facilitate tissue culture initiation. Visual assessment of micropropagated plants in the field did not show any off-types, based on gross morphological changes of plant morphology or disease reaction, compared to plants of the same genotype derived from a traditional propagation method (stem cuttings). This is the first report of energy cane and Miscanthus spp. micropropagation using the SETIS bioreactor.